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Wheat egg in vitro fusion with wheat and green bristlegrass sperm

Published online by Cambridge University Press:  20 May 2019

Dong Xiao Li ,
Hai Yan Hu ,
Zhen Gang Ru  and
Hui Qiao Tian Open the ORCID record for Hui Qiao Tian [Opens in a new window]
Dong Xiao Li
Affiliation:
Henan Institute of Science and Technology, Xinxiang 453003, China
Hai Yan Hu
Affiliation:
Henan Institute of Science and Technology, Xinxiang 453003, China Collaborative Innovation Center of Modern Biological Breeding, Henan Province
Zhen Gang Ru
Affiliation:
Henan Institute of Science and Technology, Xinxiang 453003, China Collaborative Innovation Center of Modern Biological Breeding, Henan Province
Hui Qiao Tian*
Affiliation:
School of Life Sciences, Xiamen University, Xiamen, 361005, China
*
*Address for correspondence: Hui Qiao Tian. School of Life Sciences, Xiamen University, Xiamen, Fujian 361102, China. Tel: +011 86 592 2186486. E-mail: [email protected]
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Summary

Isolated gametes can be used to investigate fertilization mechanisms, and probe distant hybridization between different species. Pollen grains of wheat and Setaria viridis are tricellular, containing sperm cells at anthesis. Sperm from these plants were isolated by breaking open pollen grains in a osmotic solution. Wheat ovules were digested in an enzyme solution for 20 min, and then transferred to an isolation solution without enzymes to separate egg cells from ovules. The fusion of wheat egg cells with wheat and S. viridis sperm was conducted using an electro-fusion apparatus. Under suitable osmotic pressure (10% mannitol), calcium concentration of 0.001% (CaCl2·2H2O), and a 30–35 V alternating electric field for 15 s, egg cells and sperm adhered to each other and became arranged in a line. Electroporation of the plasma membrane of egg cells and sperm using a 300–500 V direct-current electric field (45 µs amplitude pulse) caused them to fuse.

Keywords

EggGreen bristlegrassIn vitro fusionSpermWheat
Type
Research Article
Information
Zygote , Volume 27 , Issue 3 , June 2019 , pp. 126 - 130
Copyright
© Cambridge University Press 2019 

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